Purification and biochemical characterization of a novel transglutaminase from Mythimna separata larvae (Noctuidae, Lepidoptera)

A novel transglutaminase (MsTGase) from Mythimna separata larvae was separated and purified; its biochemical property and enzymatic catalytic activities were investigated. MsTGase was obtained chromatographically by the precipitation of Sephadex G-100 gel and DEAE-Cellulose-52 ion-exchange column with 48-fold purification and a reproducible yield of approximately 12%. Molecular weight of the MsTGase was 63.5 KDa and its N-terminal amino acid sequence was GKIEEG-LVI. Michaelis constant of the MsTGase for the substrate N-CBZ-Gln-Gly was 12.83 mM with a Vmax of 7.99 U/mL. Optimum conditions for MsTGase activity were at 42 °C and pH7.5. The enzyme didn't possess metal ion at its catalytic active site; its activity could be significantly inhibited by Mg2+, but activated by Ca2+. Chlorpyrifos and spinosad showed a strong potential to increase MsTGase activity, supporting the view that MsTGase was a novel target. Moreover, the formation of intermolecular cross-links of casein and bovine serum albumin polymerized by MsTGase in the presence of DTT was observed. These findings pave the way for future studies on the physiological role of MsTGase and the potential impact of its regulation on MsTGase-associated pest management.